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. Author manuscript; available in PMC: 2019 Oct 15.
Published in final edited form as: Dev Biol. 2018 Aug 16;442(2):288–300. doi: 10.1016/j.ydbio.2018.08.005

Figure 1. PERM-2 and PERM-4 are required for eggshell integrity.

Figure 1.

(A) Schematic representations of PERM-2, PERM-4 and CBD-1 showing the location of predicted O-linked mucin attachment sites (thin vertical black lines), chitin-binding domains (red ovals), conserved regions (thick blue lines), deletion alleles, and target regions for dsRNA-mediated interference (RNAi). Chitin-binding domains 3 and 4 of CBD-1 are disrupted by the in-frame ok2913 deletion allele. (B) Left, total brood size counts defined as the number of progeny produced from a single worm over a three-day laying period. Gray bars, hatched larvae; White bars, unhatched impermeable embryos; Blue bars, unhatched embryos permeable to Nile Blue dye, with percentage of brood showing the permeable phenotype indicated. (C) Daily brood size counts and phenotype distribution, as described in (B), for the perm-2(pmn3) null and cbd-1(ok2913) strains for three successive days after initial plating of L4-stage worms. For all graphs, data were pooled from 2–3 independent experiments per condition. Control worms were either N2 wild-type worms (deletion allele experiments) or injected with him-8 dsRNA (RNAi experiments); control data were pooled since brood size means and standard errors were similar (198±16 N2 N=11 vs. 188±13 him-8 dsRNA N=15). N=number of adults, n=number of progeny. Error bars represent standard error of the mean (SEM) for total brood size. *** p<0.0001, ** p<0.001, * p<0.05 compared to controls by unpaired student’s t-test with unequal variance. (D) Widefield fluorescence images of control and RNAi-treated embryos expressing mCherry-tagged histone H2B (magenta) and a GFP-tagged plasma membrane marker (green) that were dissected into FM4–64 dye (magenta). Arrows highlight plasma membrane stained with FM4–64 dye in permeable embryos (white in the merge). Control embryos, 0% permeable; all RNAi treated embryos, 100% permeable. Multicellular embryo shown for perm-2/4(RNAi) highlights its ability to complete multiple divisions before arresting. N=number of dissected adults, n=number of embryos. Scale bars, 10um.